Annually, 5 to 7 million Americans are treated for chronic wounds that arise from diabetes, lower extremity arterial disease, lower extremity venous disease, pressure ulcers, and traumatic injuries, including burns, demonstrating the need for new advanced technologies for treatment. Advanced Plasma Products will build upon its successful Phase I results and advance the development of the Atmospheric Plasma Wound Applicator (APWA) prototypes that inactivate biofilm without causing damage to surrounding tissues, thereby healing the wound. The prototype will be tested for its wound disinfecting capabilities with in vitro wound biofilm models of different clinically significant strains and polymicrobial biofilm. Penetration studies will be performed to evaluate the depth of plasma active species in tissue and mechanistic studies will evaluate the mechanism of inactivation of microbial cells by plasma treatment. We will demonstrate the therapeutic potential of atmospheric plasma to improve wound healing by the removal of microorganisms from the biofilm of splinted excisional wounds of normal and diabetic mice and the introduction of various gaseous nitrogen species. A second generation prototype will be built and tested in a clinical setting with equine lower limb wounds. Following this two year effort, APP will initiate FDA 510K regulatory approval process.